Alkylation of Benz[a]anthracene Affects Toxicity to Early–Life Stage Zebrafish and In Vitro Aryl Hydrocarbon Receptor 2 Transactivation in a Position‐Dependent Manner

Abstract

Polycyclic aromatic hydrocarbons (PAHs) are structurally diverse organic chemicals that can have adverse effects on the health of fishes through activation of aryl hydrocarbon receptor 2 (AhR2). They are ubiquitous in the environment, but alkyl PAHs are more abundant in some environmental matrices. However, relatively little is known regarding the effects of alkylation on the toxicity of PAHs to fishes in vivo and how this relates to potency for activation of AhR2 in vitro. Therefore, the objectives of the present study were to determine the toxicity of benz[a]anthracene and three alkylated homologs representing various alkylation positions to early life stages of zebrafish (Danio rerio) and to assess the potency of each for activation of the zebrafish AhR2 in a standardized in vitro AhR transactivation assay. Exposure of embryos to each of the PAHs caused a dose‐dependent increase in mortality and malformations characteristic of AhR2 activation. Each alkyl homolog had in vivo toxicities and in vitro AhR2 activation potencies different from those of the parent PAH in a position‐dependent manner. However, there was no statistically significant linear relationship between responses measured in these assays. The results suggest a need for further investigation into the effect of alkylation on the toxicity of PAHs to fishes and greater consideration of the contribution of alkylated homologs in ecological risk assessments. Environ Toxicol Chem 2022;41:1993–2002. © 2022 SETAC

Effects of Alkylation on Potency of Benz[a]anthracene for AhR2 Transactivation in Nine Species of Freshwater Fish

“…However, ranges of species sensitivity are influenced by alkylation, including by the position(s) of the alkyl group. In addition, the present study builds on previous research showing that while alkylation has position‐dependent effects on potency, these effects are not conserved across phylogenetically diverse species of fishes (Dubiel et al, 2022; Lille‐Langøy et al, 2021). These results further support that ecological risk assessments based solely on USEPA priority PAHs underestimate risk that alkyl PAHs might pose to fish (Barron & Holder, 2003), but an average ReP to BAA for each alkyl homologue could be calculated and used to provide an order of magnitude estimate for use across species.…”

“…In the present study, RePs were used to compare the potency of 4-MBAA, 8-MBAA, and 7,12-DMBAA to BAA. Alkylation of BAA has position-dependent effects on potency for early life-stage mortality and AhR2 activation in zebrafish, with RePs for early life-stage mortality of 1.1, 5.6, and 1.4 for 4-MBAA, 8-MBAA, and 7,12-DMBAA, respectively (Dubiel et al, 2022). The RePs for AhR2 activation in zebrafish were 1.6, 6.9, and 0.8 for 4-MBAA, 8-MBAA, and 7,12-DMBAA (Table 2).…”

“…However, consistent with TCDD, the AhR2 of lake trout was the most sensitive to activation by 7,12-DMBAA, and the AhR2 of northern pike was the least sensitive to activation by each PAH and TCDD. Previous studies investigating AhR-mediated responses in cell-based assays or embryotoxicity assays reported that alkylation can have position-dependent effects on potency of PAHs within a species (Barron et al, 2004;Donald et al, 2023;Dubiel et al, 2022;Fang et al, 2022;Lille-Langøy et al, 2021;Lin et al, 2015). However, no study to date has investigated whether effects of alkylation on potency are conserved across species.…”

“…Analytical grade BAA, 4-MBAA, 8-MBAA, and 7,12-DMBAA were purchased from Millipore-Sigma. Stock solutions for BAA, 4-MBAA, 8-MBAA, and 7,12-DMBAA of 20, 5, 4, and 11 mg/ml, respectively, which is the limit of solubility of each compound, were prepared in full-strength dimethylsulfoxide (DMSO, >99.9% pure) as described previously (Dubiel et al, 2022).…”

“…For example, while chrysene did not activate the AhR2 of Atlantic cod (Gadus morhua), each alkylated homologue did with varying degrees of potency (Lille-Langøy et al, 2021). Alkylation of benz[a]anthracene (BAA) had position-dependent effects on potency for activation of the zebrafish (Danio rerio) AhR2, with similar effects on potency also being observed for embryotoxicity (Dubiel et al, 2022). Alkylation of BAA and benzo[a]pyrene (BAP) also had positiondependent effects on the concentration that induced a 20% reduction in general morphology score of zebrafish embryos (Fang et al, 2022).…”